The goal of this revised proposal is to dissect the role of brain mast cells in sleep and behavioral controls. Mast cells are a heterogeneous population of granulocytic cells of the immune system and they contain numerous mediators, such as histamine and serotonin, cytokines, chemokines, and lipid- derived factors. Mast cells localize not only in the periphery but are also resident in the brain of mammalians. Mast cells in the brain are constitutively active, releasing their contents gradually or rapidly by anaphylactic degranulation. Their activity is also increased by a wide range of stimuli including both immune and non-immune signals. Brain mast cell neuromodulation may thus be involved in various neurological and psychiatric diseases. However, the function of mast cells in the brain for the mediation of neurobehavior is largely unknown. Using Kit mutant mast cell deficient mice (KitW/KitW-v), we have preliminary results indicating that brain mast cells regulate sleep/wake and other behavioral phenotypes and that histamine from brain mast cells promote wakefulness. However, Kit mutant mice possess pleiotropic defects in pigment-forming cells, germ cells, RBC`s and mast cells, and thus may lack specificity. Dr. Kubo's lab at Tokyo University of Science, recently produced a new inducible and Kit- independent mast cell deficient, Mas-TRECK (toxin receptor knockout) mouse. Injections of diphtheria toxin (DT) selectively deplete mast cell and basophiles in Mas-TRECK mice, whereas the numbers of other hematopoietic cell populations exhibit no changes. We also anticipate brain mast cells can be specifically depleted in these Mas-TRECK mice, if DT is administered intracerebroventricularly. Although a series of experimental evidence has suggested that neuroimmune interaction is important for sleep and neurobehavior control and for some psychiatric disorders, the roles of mast cells in the control of sleep and other behaviors has not yet been systematically evaluated. Our results showed for the first time that brain mast cells are likely involved in physiological wakefulness and arousal responses to various behavioral manipulations. We also found that the availability of mast cells influences prevalent psychiatric symptoms, including anxiety and depression-like symptoms. It is therefore conceivable that brain mast cells play a significant role in the pathophysiology of some neuropsychiatric diseases. In this revised proposal, we will therefore further examine the role of brain mast cells in sleep/wake and behavioral control in physiological and pathological conditions using Mas-TRECK mice. The knowledge obtained from the proposed experiments will likely bring a new concept about how sleep and behavior is regulated by non-neuronal cells in normal and pathophysiological conditions. Since known chemical and substances released from the mast cell is likely involved in the mediation of the effects, these systems can be targeted by pharmacological treatments for various sleep and behavioral disorders, and results from the proposed study will be very useful for further translation research.